Membrane requirement for folding of the herpes simplex virus 1 gB cytodomain suggests a unique mechanism of fusion regulation

Herpes simplex virus type 1 (HSV-1) enters cells by fusion of its envelope with a host cell membrane, which requires four viral glycoproteins and a cellular receptor. Viral fusion glycoprotein B (gB) mediates membrane fusion through the action of its ectodomain, while its cytoplasmic domain (cytodomain) regulates fusion from the opposite face of the membrane by an un-knownmechanism. The gB cytodomain appears to restrict fusion, because point or truncationmutations within it increase the extent of fusion (synmutations). Previously, we showed that the hyperfusion phenotype correlated with reducedmembrane binding in gB syn truncationmutants and proposed that membrane binding was important in regulating fusion. Here, we ex-tended our analysis to three syn point mutants: A855V, R858H, and A874P. These mutations produce local conformational changes, with some affecting membrane interaction, which suggests that while synmutants may deregulate fusion by somewhat different mechanisms, maintaining the wild-type (WT) conformation is critical for fusion regulation. We further show that the presence of a membrane is necessary for the cytodomain to achieve its fully folded conformation and propose that the mem-brane-bound form of the cytodomain represents its native conformation. Taken together, our data suggest that the cytodomain of gB regulates fusion by a novel mechanism in which membrane interaction plays a key role. Herpes simplex virus type 1 (HSV-1) causes diseases rangingfrommild skin lesions to fatal encephalitis and neonatal her-pes (28). Like all herpesviruses, HSV-1 enters host cells by fusion of the viral envelope with the plasma membrane or an endocyti